PROJECT 1 Modulation of GABA-A or NMDA receptors underlies the anesthetic actions of most clinically used general anesthetics. In the central nervous system, enhancement of GABA-A receptor function increases inhibitory neurotransmission and inhibition of NMDA receptor function decreases excitatory neurotransmission. Anesthetic steroids enhance the actions of GABA at GABA-A receptors. Steroids containing negatively charged groups inhibit GABAergic neurotransmission. NMDA receptors are modulated, either positively or negatively, by steroids that contain negatively charged groups. Thus, understanding the actions of steroids at these receptors is important for understanding the clinical effects of the steroid class of anesthetics. The three specific aims of this project are focused on gaining new knowledge of the molecular details of the interactions of steroids with GABA-A and NMDA receptors. Synthetic chemistry will be performed to prepare novel analogues of anesthetic steroids for structure-activity relationships (SAR) studies. Electrophysiological, binding, and behavioral assayswill be used for evaluation of the compounds. Specific aim 1 addresses the importance of the steroid ring system for anesthetic steroid effects at GABA-A and NMDA receptors. In specific aim 2, anesthetic steroid analogues containing fluorescent groups will be prepared to: 1) investigate how steroids access their binding sites in the transmembrane domains of GABA-A receptors;and 2) investigate the entry and distribution of anesthetic steroids in brain cells. Specific aim 3 will investigate the molecular basis for the enantioselectivity of anesthetic steroid action at GABA-A receptors. The long term goals of this project are to use the methods of medicinal chemistry to obtain new pharmacological tools for investigationof the effects of steroids at GABA-A and NMDA receptors. Ultimately, this information could lead to the discovery of new anesthetic, anticonvulsant, anxiolytic and sedative hypnotic drugs.